1. Technical Field
The present invention relates the general field of biological assays, and more particularly to a microfluidic apparatus for conducting cellular, enzymatic, chemical and molecular biological processes on a solid substrate, generally a glass slide.
2. Description of the Related Art
Detection of biomarkers at the point of care (such as, for example, in the field, in remote areas, in a doctor's office, and at the bedside in a hospital) has the potential to offer real time diagnostic information, improve patient care outcomes, decrease sample volumes, and provide analytical information from a broad range of biological samples, many of which may be acquired relatively non-invasively.
Co-assigned patents and patent applications relevant to the development of clinical assays in a microfluidic device test format include PCT Publication No. WO200201184 (“Fluid Mixing in Microfluidic Structures”), U.S. Pat. No. 6,743,399 (“Pumpless Microfluidics”), U.S. Pat. No. 6,488,896 (“Microfluidic Analysis Cartridge”), U.S. Patent Application Publication No. 20050106066 (“Microfluidic Devices for Fluid Manipulation and Analysis”), U.S. Patent Application Publication No. 20020160518 (“Microfluidic Sedimentation”), U.S. Patent Application Publication No. 20030124619 (“Microscale Diffusion Immunoassay”), U.S. Patent Application Publication No. 20030175990 (“Microfluidic Channel Network Device”), U.S. Patent Application Publication No. 20050013732 (“Method and System for Microfluidic Manipulation, Amplification and Analysis of Fluids, For Example, Bacteria Assays and Antiglobulin Testing”), U.S. Pat. No. 6,581,899 (“Valve for Use in Microfluidic Structures”), and PCT Publication No. WO2007/064635 (“Microfluidic Cell Capture and Mixing Circuit”), all of which are hereby incorporated by reference in their entireties. Also incorporated herein by reference is U.S. Pat. No. 6,729,352, which relates to microfluidic valve structures.
Capillary action has proven useful in designing small disposable diagnostic devices, as discussed in, for example, U.S. Pat. No. 5,415,994, U.S. Pat. No. 5,658,723 and PCT Publication No. WO199633399. However, to improve sensitivity, mixing during affinity capture is likely to be helpful. Mixing small volumes, however, is not without unique problems. The problem of mixing in a microvolume is variously addressed in, for example, U.S. Pat. Nos. 6,468,807, 6,916,113, 6,872,566 and 6,729,352 (which describes a “slit mixer”) and also in Hardt, S. et al. (“Passive Micromixers for Applications in the Microreactor and μTAS Fields”, Microfluid Nanofluid, vol. 1:108-118, 2005).
Further related art includes the Maui® Mixer sold by BioMicro Systems and described in PCT Publication No. WO2003015923, U.S. Patent Application Publication No. 20050019898 and U.S. Pat. No. 7,223,363. The teaching of these disclosures relates particularly to use of a pair of flexible bladders mounted at each end of a rectilinear microfluidic chamber and a gasketed assembly for sealing the chamber to a glass slide. Preferred dimensions are given, and the claims are generally directed to a rectilinear chamber with parallel sides. In particular, the height of the chamber is generally 10 to 500 μm and the height is small relative to the length and to the width. The walls of the chamber are selected to be smooth and to run parallel to the axis of flow so as to avoid trapping of air bubbles and reduced mixing efficiency. U.S. Pat. Nos. 5,100,626 and 6,303,389 also teach parallel channel walls.
Mixing is achieved with sonication in U.S. Pat. No. 6,326,211 and with agitation in U.S. Pat. No. 6,309,875. See also PCT Publication Nos. WO200201184 and WO200170381, and U.S. Pat. Nos. 6,287,850, 6,272,939, 6,158,712, 5,922,591 and 5,639,428. However, these methods depend on relatively large sample volumes, large hybridization chambers and inconvenient or complicated equipment not readily adapted to the point of care. Also of related interest, U.S. Pat. No. 5,718,567 describes a microscale diaphragm pump with check valves and a titanium diaphragm, U.S. Pat. No. 7,052,594 to Pelrine describes an electrically active diaphragm for use in microfluidic pumps, and U.S. Pat. No. 6,843,263 to Koh describes microfluidic cards with “a deformable chamber” having an elastic thin film cover and a mechanical actuator, the film serving to seal the body and the mechanical actuator serving to deform the film and move plugs of fluid in the body, relying on the elasticity of the film and open venting to generate reciprocal flow of the fluid plug. The pressurization of an open vented system containing hazardous sample is a serious disadvantage of the teachings of U.S. Pat. No. 5,718,567.
However, in contemplating use of disposable microfluidic device-based assays of clinical specimens, design of fully closed, “single-entry” systems has not been adequately addressed. In view of contamination hazards associated with working with potentially infectious human samples, resealable entry into the device with gasketed sealing means is often simply not acceptable. Moreover, the assay format must be robust and readily adapted for use with a wide range of biomarkers, both in automated and manual assays, at the point of care. To achieve these objectives, further improvement in mixing arts for microvolumes is needed.
Also needed are devices compatible with solid planar substrates such as glass slides, which are frequently used to mount, stain and examine tissue specimens, cell specimens, and to screen DNA and protein arrays, while not limited thereto, where there is a need to promote mixing in a thin fluid layer over the specimens which may have a microfluidic dimension.
Accordingly, although there have been advances in the field, there remains a need in the art for microfluidic devices that meet the foregoing criteria. The present invention addresses these needs and provides further related advantages.